Many adhesive proteins, including fibrinogen, fibronectin, von Willebrand factor, and vitronectin, contain an amino acid sequence that functions as a recognition site for adhesion receptors. The amino acid sequences within these proteins are recognized by structurally-related receptors on a variety of mammalian cells that have been termed integrins or cytoadhesions. Hynes, Cell, 48, 549-554 (1987); Ruoslahti et al., Science, 238, 491-497 (1987).
One well-characterized member of the integrin family is the blood platelet membrane glycoprotein IIb-IIIa complex (GP IIb-IIIa). Phillips et al., Blood, 71, 831-843 (1988). Upon platelet activation, GP IIb-IIIa becomes competent to bind fibrinogen, a process required for platelet aggregation. Bennett et al., J. Clin. Invest., 64, 1393-1400 (1979); Marguerie et al., J. Biol. Chem., 254, 5357-5363 (1979). Based on studies with synthetic peptides, the Arg-Gly-Asp-Ser sequence at positions 572-575 of the fibrinogen A.alpha. chain appears to play a major role in the interaction of this protein with GP IIb-IIIa, although other regions in fibrinogen may be involved as well. Gartner et al., J. Biol. Chem., 260, 11891-11894 (1985); Ginsberg et al., J. Biol. Chem., 260, 3931-3936 (1985); Haverstick et al., Blood, 66, 946-952 (1985); Pytela et al., Science, 231, 1159-1162 (1986); Kloczewiak et al., Thrombosis Res., 29, 249-255 (1983); Kloczewiak et al., Biochemistry,. 23, 1767-1774 (1984). However, the precise details of the molecular interaction between fibrinogen and GP IIb-IIIa are, at present, unknown. The tripeptide sequence Arg-Gly-Asp has also been identified in the adhesive proteins fibronectin, von Willebrand factor and vitronectin. Hynes, Cell, 48, 549-554 (1987); Ruoslahti et al., Science, 238, 491-497 (1987). A similar region of fibronectin containing the sequence Arg-Glu-Asp-Val has been implicated in the binding of this protein to melanoma cells. Humphries et al., J. Cell Biol., 103, 2637-2647 (1986). Moreover, von Willebrand factor, which can bind to GP IIb-IIIa on activated platelets, contains, in its long (significantly greater than 50 residue) amino acid sequence, an Arg-Tyr-Asp-Ala, in addition to an Arg-Gly-Asp-Ser, sequence. Titani et al., Biochemistry, 25, 3171-3184 (1986). Furthermore, the sequences Arg-Phe-Asp-Ser and Arg-Tyr-Asp-Ser are found within the long chain peptides that comprise the major histocompatibility antigens, peptides which have a length greatly in excess of 50 amino acid residues. Auffray et al., Human Immunol., 15, 381-390 (1986).
Numerous applications have been found for peptides containing the amino acid sequence Arg-Gly-Asp. The binding of fibrinogen to blood platelets, for example, has been found to be inhibited by Arg-Gly-Asp-containing peptides. In addition, PAC1, an IgM-.kappa. murine monoclonal antibody that, like fibrinogen, appears to bind to GP IIb-IIIa on activated platelets, is similarly inhibited by such peptides. The inhibition of fibrinogen and PAC1 occurs at an apparent Ki of about 10-20 .mu.M when the tetrapeptide Arg-Gly-Asp-Ser is employed. Shattil et al., J. Biol. Chem., 260, 11107-11114 (1985); Bennett et al., J. Biol. Chem., (1988); Shattil et al., Blood, 68, 1224-1231 (1986). Moreover, U.S. Pat. No. 4,683,291 discloses the usefulness of peptides containing the Arg-Gly-Asp sequence in inhibiting cellular adhesion in general, and platelet to platelet binding specifically. Such peptides are also reported as useful in retarding the formation of blood clots.
Additional and/or better peptides which meet the important ends of inhibiting fibrinogen to platelet binding, platelet to platelet binding and/or blood clot formation are needed. The present invention is directed to these and other important needs.